PROJECT SUMMARY- PROJECT 3: ESTABLISHING CLINICAL UTILITY OF CSF BIOMARKERS FOR PD Accurate diagnosis of Parkinson's disease (PD) and monitoring of PD patients remain challenging and preclude the most effective patient care. Although the NINDS and other PD patient advocacy groups such as the Michael J. Fox Foundation have recognized the need for both diagnostic and prognostic PD biomarkers, no such biomarkers have been validated thus far. This Clinical Research Project will utilize Multiple Reaction Monitoring-Mass Spectrometry (MRM-MS) to determine if specific protein phosphorylation events can be used as novel PD-specific diagnostic/prognostic biomarkers. Previous research has shown that the non-receptor tyrosine kinase, c-Abl, phosphorylates ??Synuclein at the Y-39 site and Parkin at the Y-143 site. During PD, dysregulation of these two pathways ultimately leads to neuronal cell death, which in turn leads to the clinical manifestations of PD. In preliminary studies, we have already developed MRM-MS based quantitative assays to monitor these phosphorylated and unphosphorylated forms of ?-Synuclein and Parkin in CSF samples. Our hypothesis is that dysregulation of c- Abl signaling cascade is intrinsically linked to PD pathogenesis and that the relative phosphorylation state of downstream c-Abl substrates, ?-Synuclein and Parkin, should provide a sensitive read-out for the presence and/or the severity of PD. Aim 1 will measure the relative concentration of the Y-39 tryptic peptide of ?-Synuclein in both its phosphorylated and unphosphorylated forms in the CSF of PD patients and controls. This assay will be employed in several clinical cohorts to determine whether the phosphorylated Y-39 residue can function as (i) a diagnostic marker by differentiating between PD patients and controls and/or (ii) a prognostic PD marker in patients that can be used for assessment of early and late (more severe) stages of PD. Aim 2 will adopt the same approach as described in Aim 1 to test whether c-Abl mediated phosphorylation of Parkin at Y-143 can be used to facilitate either diagnosis with PD or patient monitoring as a function of disease severity. Taken together, this project will identify and validate c-Abl substrates as novel diagnostic/prognostic protein-based biomarkers that have the potential to provide clinicians with novel strategies for establishing a definitive diagnosis of PD and/or for monitoring the severity of disease in patients with PD.